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BACKGROUND AND GENERAL INTRODUCTION 13

CHAPTER I. Effects of 5,7-DHT and DSP-4 on Nociceptive Behaviors Induced by Formalin, Substance P, or Glutamate in Mice 19

INTRODUCTION 20

MATERIALS AND METHODS 22

1. Experimental animals 22

2. Intrathecal (i.t.) injection 22

3. Formalin-induced nociceptive behavioral test 23

4. SP-and glutamate-induced nociceptive behavioral tests 23

5. Pretreatment of monoaminergic neurotoxins 23

6. Drugs 24

7. Statistical analysis 24

RESULTS 25

1. The effects of 5,7-DHT and DSP-4 on the intraplantar formalin-induced nociceptive behaviors 25

2. The effects of 5,7-DHT and DSP-4 on the intrathecal SP-induced nociceptive behaviors 25

3. The effects of 5,7-DHT and DSP-4 on the intrathecal glutamate-induced nociceptive behaviors 25

DISCUSSION 32

CHAPTER II. Effects of Methysergide on the Nociceptive Responses in Various Pain Models 34

INTRODUCTION 35

MATERIALS AND METHODS 37

1. Experimental animals 37

2. Intraperitoneal (i.p.) and intrathecal (i.t.) injections 37

3. SP-and EAAs-induced nociceptive behavioral tests 38

4. Formalin-induced nociceptive behavioral test 38

5. Drugs 39

6. Statistical analysis 40

RESULTS 41

1. The effect of methysergide on SP-induced nociceptive behaviors 41

2. The effect of methysergide on EAAs-induced nociceptive behaviors 41

3. The effect of methysergide on the formalin-induced nociceptive behaviors 41

4. The effect of 5,7-DHT or DSP-4 on the behavioral responses elicited by methysergide in the formalin test 42

DISCUSSION 51

CHAPTER III. Differential Roles of Spinal Cholera Toxin-and Pertussis Toxin-Sensitive G-Proteins in Various Pain Models 56

INTRODUCTION 57

MATERIALS AND METHODS 60

1. Experimental animals 60

2. Intrathecal (i.t.) injection 60

3. Formalin-induced nociceptive behavioral test 61

4. Capsaicin-induced nociceptive behavioral test 61

5. SP-induced nociceptive behavioral test 62

6. EAAs-induced nociceptive behavioral tests 62

7. Drugs 63

8. Statistical analysis 63

RESULTS 64

1. The effect of intrathecal (i.t.) CTX or PTX pretreatment on the nociceptive behaviors elicited by s.c. injection of formalin 64

2. The effect of i.t. CTX or PTX pretreatment on the behavioral nociceptive responses elicited by i.t. injection of capsaicin 64

3. The effect of i.t. CTX or PTX pretreatment on the behavioral nociceptive responses elicited by i.t. injection of SP 65

4. The effect of i.t. CTX or PTX pretreatment on the behavioral nociceptive responses elicited by i.t. injection of EAAs 65

DISCUSSION 75

CHAPTER IV. Involvement of Phosphorylated Ca?/Calmodulin-Dependent Protein Kinase II in the Mouse Formalin and Substance P Pain Models 79

INTRODUCTION 80

MATERIALS AND METHODS 83

1. Experimental animals 83

2. Intrathecal (i.t.) and intracerebroventricular (i.c.v.) injection of drugs 83

3. Formalin treatment and nociceptive behavioral analysis 84

4. Substance P treatment and nociceptive behavioral analysis 85

5. Western immunoblot analysis 85

6. Immunocytochemistry 86

7. Statistical analysis 87

RESULTS 88

1. The effect of intraplantar formalin on the pCaMK-II level in the spinal cord 88

2. The effect of KN-93 pretreated i.t. on the intraplantar formalin-induced nociceptive behaviors 88

3. The effect of SP injected i.t. on the pCaMK-II level in the spinal cord 88

4. The effect of KN-93 pretreated i.t. on the i.t. SP-induced nociceptive behaviors 89

5. The effect of intraplantar formalin on the pCaMK-II level in the hippocampus 89

6. The effect of intraplantar formalin on the pCaMK-II level in the hypothalamus 89

7. The effect of KN-93 pretreated i.c.v. on the intraplantar formalininduced nociceptive behaviors 90

8. The effect of SP injected i.t. on the pCaMK-II level in the hippocampus 90

9. The effect of SP injected i.t. on the pCaMK-II level in the hypothalamus 91

10. The effect of SP injected i.t. on the pCaMK-II level in the brainstem 91

11. The effect of KN-93 pretreated i.c.v. on the i.t. SP-induced nociceptive behaviors 91

DISCUSSION 103

CHAPTER V. Involvement of Phosphorylated Extracellular Signal-Regulated Kinase in the Mouse Formalin and Substance P Pain Models 106

INTRODUCTION 107

MATERIALS AND METHODS 109

1. Experimental animals 109

2. Intrathecal (i.t.) and intracerebroventricular (i.c.v.) injection of drugs 109

3. Formalin treatment and nociceptive behavioral analysis 110

4. Substance P treatment and nociceptive behavioral analysis 111

5. Western immonoblot analysis 111

6. Immunocytochemistry 112

RESULTS 114

1. The effect of intraplantar formalin on the pERK level in the spinal cord 114

2. The effect of PD98059 pretreated i.t. on the intraplantar formalininduced nociceptive behaviors 114

3. The effect of SP injected i.t. on the pERK level in the spinal cord 114

4. The effect of PD98059 pretreated i.t. on the i.t. SP-induced nociceptive behaviors 115

5. The effect of intraplantar formalin on the pERK level in the hippocampus 115

6. The effect of intraplantar formalin on the pERK level in the hypothalamus 115

7. The effect of PD98059 pretreated i.c.v. on the intraplantar formalininduced nociceptive behaviors 116

8. The effect of SP injected i.t. on the pERK level in the hippocampus 116

9. The effect of SP injected i.t. on the pERK level in the hypothalamus 116

10. The effect of SP injected i.t. on the pERK level in the brainstem 117

11. The effect of PD98059 pretreated i.c.v. on the i.t. SP-induced nociceptive behaviors 117

DISCUSSION 129

CHAPTER VI. Alterations of c-fos mRNA Expression in Hypothalamic-Pituitary-Adrenal Axis and Various Brain Regions Induced by Intrathecal Single and Repeated Substance P Administrations in Mice 132

INTRODUCTION 133

MATERIALS AND METHODS 135

1. Experimental animals 135

2. Intrathecal (i.t.) injection of substance P (SP) 135

3. Total RNA isolation and Northern blot analysis 136

RESULTS 138

1. The effect of i.t. SP treatment on the c-fos mRNA expression in the spinal cord 138

2. The effect of i.t. SP treatment on the c-fos mRNA expression in the hippocampus 138

3. The effect of i.t. SP treatment on the c-fos mRNA expression in the HPA axis 139

DISCUSSION 144

CHAPTER VII. Role of Spinal c-Jun in the Regulation of Prodynorphin mRNA Expression in Intraplantar Formalin Injected Rat 146

INTRODUCTION 147

MATERIALS AND METHODS 149

1. Treatment of animals 149

2. Isolation of total RNA and proteins 149

3. Northern blot analysis 150

4. Western blot analysis 151

5. Electrophoretic mobility shift assay 151

RESULTS 153

1. The effect of intraplantar formalin on the levels of proDYN and proENK mRNA in the spinal cord 153

2. The effect of intraplantar formalin on the levels of proto-oncoproteins in the spinal cord 153

3. The effect of intraplantar formalin on the levels of AP-1 or DYNCRE- 3DNA binding activity in the spinal cord 153

DISCUSSION 160

CONCLUSIONS 165

REFERENCES 169

ABSTRACT IN KOREAN 193

Fig. 1-1 The effects of 5,7-DHT and DSP-4 on the behavioral response elicited by the intraplantar formalin 27

Fig. 1-2 The effect of 5,7-DHT on the behavioral response elicited by the intrathecal injection of substance P 28

Fig. 1-3 The effect of DSP-4 on the behavioral response elicited by the intrathecal injection of substance P 29

Fig. 1-4 The effect of 5,7-DHT on the behavioral response elicited by the intrathecal injection of glutamate 30

Fig. 1-5 The effect of DSP-4 on the behavioral response elicited by the intrathecal injection of glutamate 31

Fig. 2-1 The effect methysergide on the nociceptive behaviors elicited by intrathecal injection of substance P in mice 43

Fig. 2-2 The effect of methysergide on nociceptive behavior induced by glutamate administered intrathecally 44

Fig. 2-3 The effect of methysergide on nociceptive behavior induced by NMDA administered intrathecally 45

Fig. 2-4 The effect of methysergide on nociceptive behavior induced by AMPA administered intrathecally 46

Fig. 2-5 The effect of methysergide on nociceptive behavior induced by kainic acid administered intrathecally 47

Fig. 2-6 The effect of methysergide on nociceptive behavior elicited by subcutaneous injection of formalin in mice 48

Fig. 2-7 The effect of 5,7-DHT on the behavioral response elicited by methysergide during the first (A) and second (B) phases of the formalin test 49

Fig. 2-8 The effect of DSP-4 on the behavioral response elicited by methysergide during the (A) first and (B) second phases of the formalin test 50

Fig. 3-1 The effect of cholera toxin (CTX) on the nociceptive behaviors elicited by subcutaneous injection of formalin in mice 67

Fig. 3-2 The effect of pertussis toxin (PTX) on the nociceptive behaviors elicited by subcutaneous injection of formalin in mice 68

Fig. 3-3 The effects of cholera toxin (CTX; A) and pertussis toxin (PTX; B) on the nociceptive behaviors elicited by capsaicin administered intrathecally 69

Fig. 3-4 The effects of cholera toxin (CTX; A) and pertussis toxin (PTX; B) on the nociceptive behaviors elicited by substance P administered intrathecally 70

Fig. 3-5 The effects of cholera toxin (CTX; A) or pertussis toxin (PTX; B) on the nociceptive behaviors elicited by glutamate administered intrathecally 71

Fig. 3-6 The effects of cholera toxin (CTX; A) or pertussis toxin (PTX; B) on nociceptive behaviors elicited by NMDA administered intrathecally 72

Fig. 3-7 The effects of cholera toxin (CTX; A) or pertussis toxin (PTX; B) on nociceptive behaviors elicited by AMPA administered intrathecally 73

Fig. 3-8 The effects of cholera toxin (CTX; A) or pertussis toxin (PTX; B) the nociceptive behaviors elicited by kainic acid administered intrathecally 74

Fig. 4-1 The effect of intraplantar injection of formalin on the pCaMKII expressions in the spinal cord 92

Fig. 4-2 The effect of KN-93 pretreated spinally on the behavioral response elicited by the intraplantar formalin 93

Fig. 4-4 The effect of KN-93 pretreated spinally on the behavioral response elicited by the intrathecal substance P 95

Fig. 4-7 The effect of KN-93 pretreated supraspinally on the behavioral response elicited by the intraplantar formalin 98

Fig. 4-8 The effect of intrathecal injection of substance P on the pCaMK-II expressions in the hippocampus 99

Fig. 4-9 The effect of intrathecal injection of substance P on the pCaMK-II expressions in the hypothalamus 100

Fig. 4-10. The effect of intrathecal injection of substance P on the pCaMK-II expressions in locus ceruleus 101

Fig. 4-11 The effect of KN-93 pretreated supraspinally on the behavioral response elicited by the intrathecal substance P 102

Fig. 5-1 The effect of intraplantar injection of formalin on the pERK expressions in the spinal cord 118

Fig. 5-2 The effect of PD98059 pretreated spinally on the behavioral response elicited by the intraplantar formalin 119

Fig. 5-3 The effect of intrathecal injection of substance P on the pERK expressions in the spinal cord 120

Fig. 5-4 The effect of PD98059 pretreated spinally on the behavioral response elicited by the intrathecal substance P 121

Fig. 5-5 The effect of intraplantar injection of formalin on the pERK expressions in the hippocampus 122

Fig. 5-6 The effect of intraplantar injection of formalin on the pERK expressions in the hypothalamus 123

Fig. 5-7 The effect of PD98059 pretreated supraspinally on the behavioral response elicited by the intraplantar formalin 124

Fig. 5-8 The effect of intrathecal injection of substance P on the pERK expressions in the hippocampus 125

Fig. 5-9 The effect of intrathecal injection of substance P on the pERK expressions in hypothalamus 126

Fig. 5-10 The effect of intrathecal injection of substance P on the pERK expressions in locus ceruleus 127

Fig. 5-11 The effect of PD98059 pretreated supraspinally on the behavioral response elicited by the intrathecal substance P 128

Fig. 6-1 The effect of a single or repeated administration of substance P (SP) on the c-fos mRNA level in the spinal cord 140

Fig. 6-2 The effect of a single or repeated administration of substance P (SP) on the c-fos mRNA level in the hippocampus 141

Fig. 6-3 The effect of a single or repeated administration of substance P (SP) on the c-fos mRNA level in the hypothalamic-pituitary-adrenal (HPA) axis 143

Fig. 7-1 The effect of intraplantar formalin on prodynorphin mRNA expression in rat spinal cord 155

Fig. 7-2 The effect of intraplantar formalin on proenkephalin mRNA expression in rat spinal cord 156

Fig. 7-3 The effect of intraplantar formalin on c-Jun protein expression in rat spinal cord 157

Fig. 7-4 The effect of intraplantar formalin on the AP-1 DNA binding activity in rat spinal cord 158

Fig. 7-5 The self-competition and antibody interference studies of intraplantar formalin induced AP-1 DNA binding activity 159